Polycomblike: A Gene That Appears to Be Required for the Normal Expression of the Bithorax and Antennapedia Gene Complexes of DROSOPHILA MELANOGASTER

A newly identified gene is described that is required for the maintenance of normal identities in many of the body segments of the fly. The effects of mutants in this gene, which is called Polycomblike (Pcl), suggest that its wild-type allele functions in the regulation of the bithorax gene complex (BX-C) and the Antennapedia gene complex (ANT-C). Evidence in favor of this idea derives from (1) the close correspondence between segmental transformations caused by Pcl mutants and those caused by dominant gain-of-function mutants in the BX-C and ANT-C, (2) the interactions observed between Pcl mutants and mutants in these complexes, and (3) the dependence upon BX-C and ANT-C dosage of the severity of at least one of the transformations caused by Pcl mutants. Arguments are presented that the control of the BX-C and ANT-C by Pcl⁺ is negative in nature. The results of clonal analysis experiments indicate that, at least for the BX-C, Pcl⁺ exerts this control until late in development. Since the wild-type allele of another gene, called Polycomb (Pc), has previously been shown to have many of the same properties as Pcl⁺, it appears that the BX-C and perhaps also the ANT-C are continuously regulated during development by at least two and probably several other genes.     

Maternal and zygotic requirement for thepolyhomeotic complex genetic locus inDrosophila

Summary The complex genetic locuspolyhomeotic (ph) is a member of thePolycomb (Pc)-group of genes and as such is required for the normal expression of ANT-C and BX-C genes. It also has probably other functions since amorphicph alleles display a cell death phenotype in the ventral epidermis of 12-h-old embryos. Here it is shown that lethal alleles ofph (amorph and strong hypomorph) show transformation of most of their segments towards AB8. Theph ⁺ product is required autonomously in imaginal cells. The total lack ofph ⁺ function prevents viability of the cuticular derivatives of these cells.ph has a strong maternal effect on segmental identity and epidermal development that can not be rescued by one paternally supplied dose ofph ⁺ in the zygote. These phenotypes differ substantially from those of previously describedPc-group genes. AmongPc-group genes,ph seems to be the only one that is strongly required both maternally and zygotically for normal embryonic development.     

Genetic interactions between the Polycomb locus and the Antennapedia and Bithorax complexes of Drosophila

Summary We have studied the embryonic and adult phenotypes of genetic combinations between Polycomb (Pc), Regulator of bithorax (Rg-bx) and the genes of the Bithorax complex (BX-C) and the Antennapedia complex (ANT-C). The products of Pc and Rg-bx genes act antagonistically, their mutant combinations leading to the ectopic expression of genes of the BX-C and ANT-C. The genetic analysis of the Pc locus suggests it is a complex gene. Pc⁺ products behave as members of a regulatory set that negatively control the expression of BX-C and ANT-C genes. Genetic combinations between different doses of Pc, Rg-bx and the genes of the BX-C and ANT-C have phenotypes which may be interpreted as resulting from ectopic derepression of posterior selector genes repressing selector genes of anterior segments. The transformation phenotypes of certain genetic combinations differ in embryos and adults. A model of regulation of the BX-C and the ANT-C genes during the imaginal cell proliferation is presented, in which the specification state is maintained by self-activation of a given selector gene and down modulation of other selector genes in the same cell.     

Head and tail development of the Drosophila embryo involves spalt, a novel homeotic gene

Mutations in spalt (sal), a novel homeotic gene on the second chromosome of Drosophila, cause opposite transformations in two subterminal regions of the embryo: posterior head segments are transformed into anterior thoracic structures and anterior tail segments are transformed into posterior abdominal structures. The embryonic phenotypes of double mutants for sal and various Antennapedia (ANT-C) or bithorax (BX-C) genes indicate that sal acts independently of the hierarchical order of the latter gene complexes. Trans-regulatory gene mutations causing ectopic expression of ANT-C and BX-C genes do not change the realms of sal action. It is proposed that the region-specific action of the sal gene primarily promotes head as opposed to trunk development, while the BX-C gene AbdB distinguishes tail from head.     

Establishment of imaginal discs and histoblast nests in Drosophila

In Drosophila the homeotic genes of the bithorax-complex (BX-C) and Antennapedia-complex (ANT-C) specify the identity of segments. Adult segment primordia are established in the embryo as the histoblast nests of the abdomen and the imaginal discs of the head, thorax and terminalia. We have used a molecular probe for the limb primordia and in vivo culture to describe the nature of the adult primordia in mutants in which the pattern of homeotic gene expression was altered. The results suggest that the histoblast or disc 'mode' of development is initiated by the extended germ band stage through activity of the BX-C and ANT-C and is relatively inflexible thereafter [corrected].     

A pupal lethal mutation with a paternally influenced maternal effect on embryonic development in Drosophila melanogaster

The maternal effect and zygotic phenotype of l(1)pole hole (l(1)ph) is described. l(1)ph is a zygotic lethal mutation which affects cell division of adult precursor cells in Drosophila larvae. The locus is located in 2F6 on the salivary gland chromosome map and four alleles have been characterized. Germ-line clonal analysis of amorphic alleles indicates that l(1)ph has a maternal effect lethal phenotype. Two lethal phenotypes are observed among embryos derived from female germ-line clones homozygous for amorphic alleles dependent upon the zygotic activity of l(1)ph⁺ introduced via the sperm. Class 1: If no wild-type dose of the gene is introduced, embryos form abnormal blastoderms in which nuclear migration and cell formation is disrupted leading to an ill-defined cuticular pattern. Class 2: If a wild-type copy of the gene is introduced, blastoderm cells do not form beneath the pole cells (the pole hole phenotype); subsequently such embryos are missing cuticular structures posterior to the seventh abdominal segment (the torso phenotype). When the zygotic activity l(1)ph⁺ is modulated using position effect variegation a new phenotype is observed among class 2 embryos in which torso embryos are twisted along their longitudinal axis.     

Direct control of antennal identity by the spineless-aristapedia gene of Drosophila

Summary Loss-of-function mutations in the spineless-aristapedia gene of Drosophila (ss ^a mutants) cause transformations of the distal antenna to distal second leg, deletions or fusions of the tarsi from all three legs, a general reduction in bristle size, and sterility. Because ss ^a mutants are pleiotropic, it has been suggested that ss ⁺ has some rather general function and that the ss ^a antennal transformation is an indirect consequence of perturbations in the expression of other genes that more directly control antennal or second leg identity. Here we test whether the ss ^a transformation results from aberrant expression of Antennapedia (Antp), a homeotic gene thought to specify directly the identity of the second thoracic segment. We find that Antp ^– ss ^a mitotic recombination clones in the distal antenna behave identically to Antp ⁺ ss ^a clones, and are transformed to second leg. This demonstrates that the ss ^a antennal transformation is independent of Antp ⁺, and suggests that ss ⁺ may itself directly define distal antennal identity. The results also reveal that Antp ⁺ is not required for the development of distal second leg structures, as these develop apparently normally in Antp ^– ss ^a antennal clones. Because Antp ^– mutations cause deletions or transformations that are restricted to proximal structures, whereas ss ^a alleles cause similar defects that are distally restricted, we suggest that ss ⁺ and Antp ⁺ may play similar, but complementary, roles in the distal and proximal portions of appendages, respectively.     

Terminal versus segmental development in the Drosophila embryo: the role of the homeotic gene fork head

Summary Mutations of the homeotic gene fork head (fkh) of Drosophila transform the non-segmented terminal regions of the embryonic ectoderm into segmental derivatives: Pre-oral head structures and the foregut are replaced by post-oral head structures which are occasionally associated with thoracic structures. Posterior tail structures including the hindgut and the Malpighian tubules are replaced by post-oral head structures associated with anterior tail structures. The fkh gene shows no maternal effect and is required only during embryogenesis. The phenotypes of double mutants indicate that fkh acts independently of other homeotic genes (ANT-C, BX-C, spalt) and caudal. In addition, the fkh domains are not expanded in Polycomb (Pc) group mutant embryos. Ectopic expression of the homeotic selector genes of the ANT-C and BX-C in Pc group mutant embryos causes segmental transformations in terminal regions of the embryo only in the absence of fkh gene activity. Thus, fkh is a region-specific homeotic rather than a selector gene, which promotes terminal as opposed to segmental development. Offprint requests to: Institut für Biologie II (Genetik), Universität Tübingen, Auf der Morgenstelle 28, D-7400 Tübingen, Federal Republic of Germany     

The multi sex combs gene of Drosophila melanogaster is required for proliferation of the germline

We present a genetic analysis showing that the Drosophila melanogaster gene multi sex combs (mxc; Santamaria and Randsholt 1995) is needed for proliferation of the germline. Fertility is the feature most easily affected by weak hypomorphic mutations of this very pleiotropic locus. Pole cell formation and early steps of gonadogenesis conform to the wild-type in embryos devoid of zygotic mxc ⁺ product. mxc mutant gonad phenotypes and homozygous mxc germline clones suggest a role for mxc ⁺ in control of germ cell proliferation during the larval stages. mxc ⁺ requirement is germ cell autonomous and specific in females, whilst in males mxc ⁺ product is also needed in somatic cells of the gonads. Although mxc can be classified among the Polycomb group (Pc-G) of genes, negative trans-regulators of the ANT-C and BX-C gene complexes, germline requirement for mxc appears independent of a need for other Pc-C gene products, and mxc gonad phenotypes are different from those induced by mutations in BX-C genes. We discuss the possible functions of the mxc ⁺ product which helps to maintain homeotic genes repressed and prevents premature larval haemocyte differentiation and neoplasic overgrowth, but promotes growth and differentiation of male and female gonads.F.D. and O.S. should be considered as equal first authors     

Genetics of gliding motility in Myxococcus xanthus (Myxobacterales): Two gene systems control movement

Summary A large number of motility mutants of the gliding bacterium Myxococcus xanthus have been isolated and analyzed by transduction. Almost all nonmotile mutants are found to be double mutants. This is explained by the existence of two parallel and almost independent multi-gene systems controlling motility, in which case at least one mutation in each system would be required eliminate motility. Only one locus, called mgl, has been found to be shared by both systems. Wild type cells move singly and in groups. Single cells move if they carry a complete gene system A, the genes of which are described in the preceding paper. Groups of cells can move if they carry a complete gene system S, but single A^–S⁺ cells do not move. Linkage analysis reveals at least 9 different loci in system S. One class of S^– mutants, those mutated in the locus tgl, are conditional mutants which, after contact with tgl ⁺ cells, become temporarily motile as cell groups. Most system A mutations have little effect on fruiting but many system S mutations block it, suggesting that system S plays a role in the fruiting process.     

Mitochondrial and nuclear myxothiazol resistance in Saccharomyces cerevisiae

Summary Mitochondrial and nuclear mutants resistant to myxothiazol were isolated and characterized. The mitochondrial mutants could be assigned to two loci, myx1 and myx2, by allelism tests. The two loci map in the box region, the split gene coding for apocytochrome b. Locus myx1 maps in the first exon (box4/5) whereas myx2 maps in the last exon (box6). The nuclear mutants could be divided into three groups: two groups of recessive mutations and one of dominant mutations. Respiration of isolated mitochondria from mitochondrial mutants is resistant to myxothiazol. These studies support the conclusion that myxothiazol is an inhibitor of the respiratory chain of yeast mitochondria. The site of action of myxothiazol is mitochondrial cytochrome b.Abbreviations box mosaic gene coding for apocytochrome b - cyt b cytochrome b - MIC minimum inhibitory concentration - MNNG N-methyl-N'-nitro-N-nitrosoguanidine - Myx ^R/Myx ^S allelte forms of a locus conferring myxothiazol resistance - myx1, myx2 mitochondrial loci conferring myxothiazol resistance - rho ⁺/rho ^– grande/cytoplasmic petite - rho ⁰ cytoplasmic petite that is deleted of all mitochondrial DNA     

低H~+_-ATPase活性植物乳杆菌突变菌筛选及基因表达的相对定量分析

【目的】筛选H~+_-ATPase活性降低的植物乳杆菌突变菌,比较其与亲本菌基因表达水平的差异,进一步探索H~+_-ATPase的调控机制。【方法】利用硫酸新霉素诱变、筛选突变菌,并对亲本菌(ZUST)和突变菌(ZUST-1、ZUST-2)进行生长、产酸能力及H~+_-ATPase活性的测定。分别提取亲本菌和突变菌的基因组DNA,扩增H~+_-ATPase全部编码基因并测序。通过荧光定量PCR对H~+_-ATPase全部编码基因进行相对定量分析。【结果】突变菌的生长和产酸能力均低于亲本菌,突变菌ZUST-1和ZUST-2的H~+_-ATPase活性比亲本菌分别降低了10.1%和28.8%。突变菌ZUST-1和ZUST-2的atp A基因均有22个位点发生突变,而ZUST-2的atp C基因有6个位点发生突变。突变菌ZUST-1和ZUST-2的atp A在对数期基因表达水平分别比亲本菌ZUST下调了41.1%和35.7%,在稳定期分别下调了43.6%和14.2%;ZUST-1的atp C基因在对数期的表达水平比ZUST略高,在稳定期比ZUST上调了30%,而ZUST-2的atp C基因未表达。【结论】突变菌H~+_-ATPase活性减弱会导致其全部编码基因在稳定期表达水平上调(除ZUST-2的atp C不表达外),而且atp A和atp C基因突变导致的基因表达水平的差异是影响H~+_-ATPase活性的主要因素,此研究结果为进一步研究植物乳杆菌中H~+_-ATPase的调控机制奠定了基础。     

Preferential mutagenesis of lacZ integrated at unique sites in the Escherichia coli chromosome

To study the variation in spontaneous mutation frequencies in different chromosomal domains, a mini-Mu-kan-lacZ ⁻transposable element was constructed to insert the lacZ ⁻(Trp570 → Opal) allele into many different loci in the Escherichia coli chromosome. Papillation on MacConkey lactose plates was used to screen for mini-Mu insertion mutants with elevated levels of spontaneous mutagenesis of lacZop → LacZ⁺ candidates were then screened for normal mutation frequencies in other genes. Two different insertion mutants with this enhanced mutagenesis phenotype were isolated from 14 000 colonies, and named plm-1 (preferential lacZmutagenesis) and plm-2. The frequency of LacZ⁻→ LacZ⁺ mutations in these plm mutants was over 400-fold higher than that in isogenic strains containing mini-Mu-kan-lacZop insertions at other loci. Six Lac⁺ reversion (or suppression) mutations obtained from each of the two plm mutants were mapped by P1 transduction and all were found to be linked to the Kan^r gene in the mini-Mu-kan-lacZop, suggesting that a localized mutagenic event is responsible for the preferential mutagenesis. Furthermore, both the LacZ⁺→ LacZ⁻and Kan^r→ Kan^s mutant frequencies of these Lac⁺ revertants were in the range of 10⁻³ to 10⁻², indicating that this putative localized mutagenesis is neither allele nor gene specific. To identify the plm loci, the chromosomal regions flanking the mini-Mu insertion sites were cloned and sequenced. A computer-assisted database search of homologous sequences revealed that the plm-1 locus is identical to the mutS gene; the mini-Mu insertion most probably results in the production of a truncated MutS protein. We suggest that the enhanced lacZ mutation frequency in plm-1 may be associated with an active process involving the putative truncated MutS protein. The DNA sequence of the plm-2 locus matched a putative malate oxidoreductase gene located at 55.5 min of the E. coli chromosome. Received: 1 August 1996 / Accepted: 3 April 1997     

A gene that regulates the bithorax complex differentially in larval and adult cells of Drosophila

Loss-of-function mutations of a new homeotic gene, sxc, in Drosophila cause transformations of body segments, suggesting inappropriate expression of BX-C and ANT-C genes. I present evidence that sxc+ is required during embryogenesis for the selective repression of the BX-C in different larval segments and show that this requirement may be met entirely by maternally derived gene product. sxc+ is also required later in development to ensure the appropriate expression of ANT-C and BX-C genes in adult thoracic and abdominal segments. Absence of sxc+ in the mesothorax apparently results in the ectopic expression of the bx+ (or Ubx+) function in both the anterior and posterior compartments; this suggests that pbx mutations may define a regulatory rather than a structural function.     

Toxicity of and mutagenesis by chlorate are independent of nitrate reductase activity in Chlamydomonas reinhardtii

Summary Spontaneous chlorate-resistant (CR) mutants have been isolated from Chlamydomonas reinhardtii wildtype strains. Most of them, 244, were able to grow on nitrate minimal medium, but 23 were not. Genetic and in vivo complementation analyses of this latter group of mutants indicated that they were defective either at the regulatory locus nit-2, or at the nitrate reductase (NR) locus nit-1, or at very closely linked loci. Some of these nit-1 or nit-2 mutants were also defective in pathways not directly related to nitrate assimilation, such as those of amino acids and purines. Chlorate treatment of wild-type cells resulted in both a decrease in cell survival and an increase in mutant cells resistant to a number of different chemicals (chlorate, methylammonium, sulphanilamide, arsenate, and streptomycin). The toxic and mutagenic effects of chlorate in minimal medium were not found when cells were grown either in darkness or in the presence of ammonium, conditions under which nitrate uptake is drastically inhibited. Chlorate was also able to induce reversion of nit ^– mutants of C. reinhardtii, but failed to produce His ⁺ revertants or Ara^r mutants in the BA-13 strain of Salmonella typhimurium. In contrast, chlorate treatment induced mutagenesis in strain E1F1 of the phototrophic bacterium Rhodobacter capsulatus. Genetic analyses of nitrate reductase-deficient CR mutants of C. reinhardtii revealed two types of CR, to low (1.5 mM) and high (15 mM) chlorate concentrations. These two traits were recessive in heterozygous diploids and segregated in genetic crosses independently of each other and of the nit-1 and nit-2 loci. Three her loci and four lcr loci mediating resistance to high (HC) and low (LC) concentrations of chlorate were identified. Mutations at the nit-2 locus, and deletions of a putative locus for nitrate transport were always epistatic to mutations responsible for resistance to either LC or HC. In both nit ⁺ and nit ^– chlorate-sensitive (CS) strains, nitrate and nitrite gave protection from the toxic effect of chlorate. Our data indicate that in C. reinhardtii chlorate toxicity is primarily dependent on the nitrate transport system and independent of the existence of an active NR enzyme. At least seven loci unrelated to the nitrate assimilation pathway and mediating CR are thought to control indirectly the efficiency of the nitrate transporter for chlorate transport. In addition, chlorate appears to be a mutagen capable of inducing a wide range of mutations unrelated to the nitrate assimilation pathway.     

Conditional lethal phosphatidylserine decarboxylase mutants of Escherichia coli

Summary The final step in the biosynthesis of phosphatidylethanolamine, the major membrane lipid of Escherichia coli, is catalyzed by the membrane-bound enzyme, phosphatidylserine decarboxylase. A variation of a procedure for localized mutagenesis (Hong and Ames, 1971) was employed to generate conditional lethal mutants in phosphatidylserine decarboxylase. In our modification, an episome carrying the psd gene closely linked to purA ⁺ was heavily mutagenized in vivo in a strain also lysogenic for phage P1 CMclr100. After induction of a phage lytic cycle, the purA ⁺ marker was transduced to a purA ^- recipient. A majority of the Pur⁺ transductants thus contained a psd gene originating from the heavily mutagenized episomal strain. Three mutants were isolated in which temperature-sensitive growth is caused by thermosensitive phosphatidylserine decarboxylase activity that is defective in vivo at the non-permissive temperature. All 3 mutations were mapped at the same location as psd1, being cotransduced with melA, purA, and ampA. The gene order in this region, as determined by a phage P1-mediated, three-factor cross is ampA-psd-purA. psd ⁺ is dominant to the psd mutant alleles.     

The dominant Drop eye mutations of Drosophila melanogaster define two loci implicated in normal eye development

The three existing dominant gain-of-function Drop alleles, Dr ¹, Dr ^Mio and Dr ^We , previously assumed to define a single locus, severely disrupt eye development. Genetic analysis of ethylmethanesulphonate (EMS) and irradiation-induced revertants revealed that the Drop mutations define two loci: the Drop locus, which is defined by the Dr ¹ and Dr ^Mio mutants, and a separate locus defined by the Dr ^We mutation, which has been renamed Wedge. The majority of the Dr ¹ and Dr ^Mio revertants are embryonic lethal in trans, mutant embryos exhibiting trachea that fail to join the Filzkörper, thus revealing a role for the Drop gene in embryogenesis. Clonal analysis of lethal revertant alleles suggests a role for both genes in eye development. In the Drop homozygous mutant clones, the outer photoreceptor cells R1–R6 develop aberrantly. Wedge, however, is not required by the developing photoreceptor cells but its absence does disrupt normal ommatidial alignment. Although the Drop and nearby string loci were shown to be genetically distinct, both Dr ¹ and Dr ^Mio were found to interact in trans with lesions at the string locus, causing loss and derangement of bristles and loss of neuromuscular coordination.     

New mutations in cloned Escherichia coli umuDC genes: Novel phenotypes of strains carrying a umuC125 plasmid

The umuDC locus of Escherichia coli is required for most mutagenesis by UV and many chemicals. Mutations in E. coli umuDC genes cloned on pBR322-derived plasmids wer e isolated by two methods. First, spontaneously-arising mutant umuDC plasmids that failed to confe cold-sensitive growth on a lexA51(Def) strain were isolated by selection. Second, mutant umuDC plasmids that affected apparent mutant yield after UV-irradiation in a strain carrying umuD⁺C⁺ in the chromosome were isolated by screening hydroxylamine-mutagenized umuD⁺C⁺ plasmids. pBR322-derived umuD⁺C⁺ plasmids inhibited the induction of the SOS response of lexA⁺ strains as measured by expression of din::Mu dl(lac) Ap) fusionsbut most mutant plasmids did not. Mutant plasmids defective in complementation of chromosomal umuD44, umuC36, or both were found among those selected for failure to confer cold-sensitivity, whereas those identified by the screening procedure yielded mostly mutant plasmids with more complex phenotypes. We studied in greater detail a plasmid pLM109, carrying the umuC125 mutation. This plasmid increased the sensitivity of lexA⁺ strainsto killing by UV-irradiation but was able to complement the deficiencies of umuC mutants in UV mutagenesis. pLM109 failed to confer cold-sensitive growth on lexA(Def) strains but inhibited SOS induction in lexA⁺ strains. The effect of pLM109 on the UV sensitivity of lexA(Def)strains was similar to that of the parental umuD⁺C⁺ plasmid. The mutation responsible for the phenotypes of pLM109 was localized to a 615-bp fragment. DNA sequencing revealed that the umuC125mutation was a G:C → A:T transition that changed codon 39 of umuC from GCC → GTC thus changing Ala39 to Val39. The implications of the umuC125 mutation for umuDC-dependent effects on UV-mutagenesis and cell survival after UV damage are discussed.